In urban ecosystems, microbes play a key role in maintaining major ecological functions that directly support human health and city life. However, the knowledge about the species composition and functions involved in urban environments is still limited, which is largely due to the lack of reference genomes in metagenomic studies comprises more than half of unclassified reads. Here we uncovered 732 novel bacterial species from 4728 samples collected from various common surface with the matching materials in the mass transit system across 60 cities by the MetaSUB Consortium. The number of novel species is significantly and positively correlated with the city population, and more novel species can be identified in the skin-associated samples. The in-depth analysis of the new gene catalog showed that the functional terms have a significant geographical distinguishability. Moreover, we revealed that more biosynthetic gene clusters (BGCs) can be found in novel species. The co-occurrence relationship between BGCs and genera and the geographical specificity of BGCs can also provide us more information for the synthesis pathways of natural products. Expanded the known urban microbiome diversity and suggested additional mechanisms for taxonomic and functional characterization of the urban microbiome. Considering the great impact of urban microbiomes on human life, our study can also facilitate the microbial interaction analysis between human and urban environment.